Intranasal immunization of cocktail/fusion protein containing Tir along with ΔG active fragment of Zot as mucosal adjuvant confers enhanced immunogenicity and reduces E. coli O157:H7 shedding in mice.

ABSTRACT

Ruminants are the major reservoirs of Escherichia coli O157H7 and its fecal shedding mainly act as a source of entry of this pathogen into the human food chain. In humans, E. coli O157H7 infection causes diarrhea, hemorrhagic colitis and hemolytic uremic syndrome. Intimate adherence of E. coli O157H7 is mediated by Translocated intimin receptor (Tir) to which intimin binds in the host cell. Since E. coli O157H7 colonizes intestinal epithelium, the mucosal vaccine has a potential to prevent its colonization. Zonula occludens toxin (Zot) of Vibrio cholerae transiently, reversibly alters epithelial tight junction structure to increase mucosal permeability of macromolecules via paracellular route. The C-terminal region of Zot (ΔG) responsible for this function could be used for mucosal antigen delivery. Therefore, we employed individual (Tir), cocktail (ΔG + Tir), fusion protein (ΔG-Tir) and assessed the efficacy of its intranasal immunization on immunogenicity and fecal shedding of E. coli O157H7 in streptomycin treated mouse model. Compared to control, ΔG + Tir, ΔG-Tir immunized mice elicited significant antigen specific antibody titers in serum (IgG, IgA) and feces (IgA), whereas Tir immunized mice induced only serum IgG titer. Cytokine analysis revealed mixed Th1/Th2 type immune response in case of ΔG + Tir, ΔG-Tir group while that of Tir group was solely Th2 type. Tir, ΔG + Tir and ΔG-Tir immunized mice showed reduction in shedding of E. coli O157H7 compared to control group. However, ΔG-Tir immunized group performed better than ΔG + Tir, Tir group in reducing fecal shedding. Overall, our results demonstrate that intranasal immunization of ΔG-Tir induces effective systemic, mucosal, cellular immune responses and represents a promising mucosal subunit vaccine to prevent E. coli O157H7 colonization.